We propose to isolate, purify and characterize the endogenous nuclease from rabbit thymus nuclei responsible for the production of the salt-soluble chromatin studied in this laboratory. The activity of this enzyme towards chromatin and DNA will be compared. Linear, circular and superhelical DNAs will also be used as substrates to gain an idea as to the types of DNA configuration the enzyme prefers. The soluble chromatin and nucleosome sub-particles will be examined at various ionic strengths and conformational changes probed using circular dichroism and electron microscopy. A study of the nucleosome containing 200 b.p. of DNA and with or without histone H1 will be aimed at elucidation of the location and role of H1 in chromatin structure. We propose to demonstrate that H1 is not combined with the linker DNA between sub-units. RNA polymerase II from rabbit thymus nuclei will be isolated and studied using both chromatin and the nucleosome as a template. A light chromatin fraction which binds RNA polymerase but does not initiate will be prepared and this will be cmpared with a non-binding fraction of chromatin. This will involve protein assays with gel electrophoresis, examination of CD spectra and measurement of rates of digestion with micrococcal nuclease and DNase I. Activation of the binding type of chromatin for RNA sysnthesis will be attempted by removal of histone H1, by acetylation of the histones or by tryptic digestion of the ends of the histones. Various nucleosomal sub-units will also be tested as templates for RNA synthesis.